Induction of endogenous tumor necrosis factor by OK-432 in ovarian cancer patients with ascites

To four ovarian cancer patients with malignant ascites, 10 KE of OK-432 was intraperitoneally administered four times at 2 day intervals for priming, and 40 KE of OK-432 was given on the 13th day after the first injection for triggering. The changes in blood monocyte and peritoneal macrophage levels and the production of tumor necrosis factor (TNF) by blood mononuclear cells (BMCs) and ascitic lymphoid cells (ALCs) were examined. In the two patients in whom TNF was induced in the ascites, TNF production by BMCs and ALCs was noted during priming. After triggering, increases in both the number of peritoneal macrophages and TNF production by ALCs were noted. In the other two patients, in whom TNF was not detected in the ascites, the ratio of peritoneal macrophages to ALCs did not change throughout the study period, and TNF production by the ALCs was not augmented. These findings suggest that OK-432 can exert a primary effect on both peritoneal macrophages and blood monocytes, and that OK-432 triggering can promote an increase in primed peritoneal macrophages and the release of TNF from these cells.     

Induction of tumor growth inhibitory factor (TGIF) in human mononuclear cells by OK-432, a streptococcal preparation

Summary A tumor growth inhibitory factor (TGIF) was induced in the culture supernatant from mixed culture of human peripheral blood mononuclear cells (PBMC) and a streptococcal preparation, OK-432, in vitro. The activity generated in the supernatant increased in a time-dependent fashion and first appeared 6 h after the initiation of culture, reaching its maximum around 48 h. The TGIF was cytostatic against seven of ten human tumor targets, but not against three murine tumor targets. Tumor cell growth was inhibited by a transient contact, i.e., 1 h, with TGIF. The TGIF was produced by lymphocytes but not by monocytes, because the activity was usually enhanced by elimination of plastic-adherent cells from the original PBMC fraction. The TGIF was relatively stable against heating at 56° C for 30 min, but the activity was totally destroyed after heating at 70° C for 5 min. The molecular weight of TGIF was estimated to be about 43×10³ daltons by gel filtration. No interferon (IFN) activity was detected in the TGIF-positive fractions obtained by gel filtration, and the TGIF-positive fractions did not inhibit the growth of tumor necrosis factor (TNF)-sensitive mouse L929 cells. The TGIF activity was not significantly affected in neutralizing tests using specific antibodies against human IFN and TNF. The OK-432 was administered i.p. for management of cancer patients with malignant ascites. Ascites-derived mononuclear cells (ASMC) were obtained before and 3 to 5 days after OK-432 injection. The ASMC obtained after the injection produced TGIF in vitro in the absence of OK-432; the preinjection ASMC showed no such production. A positive correlation was found between TGIF-producing activity by ASMC and the effect of OK-432 injection on ascites volume. These results indicate that TGIF is induced in mononuclear cells by OK-432 not only in vitro but also in vivo and plays an important role in inhibition of tumor growth in cancer patients.     

Induction of antitumor L3T4-positive T cells by OK-432 at tumor sites in mice

We have previously reported the development of antitumor effector cells by day 12 after tumor implantation using a murine malignant ascites model with BAMC-1 tumor, which could be cured completely by five consecutive i.p. injections of OK-432 starting on day 2. In contrast, the OK-432 treatment with the same protocol failed to cure the tumor-bearing athymic mice, though it could suppress tumor growth temporarily. The results suggest that T cells may play a critical role in achieving a therapeutic effect. The present study was designed to clarify the nature of the antitumor effector cells induced by OK-432 in euthymic mice. The number of tumor cells in the pertioneal cavity of OK-432-treated euthymic mice increased gradually up to day 12 and dropped suddenly on day 14, while in the athymic mice the tumor cells transiently decreased in the first 7 days then started to expand drastically on day 8. The timing of the appearance of the effector cells was examined by adoptive-transfer experiments. The peritoneal exudate cells (PEC) obtained from BAMC-1 bearing euthymic mice on various days during the treatments with OK-432 were passively transferred intraperitoneally on the respective days (synchronous transfer) or on day 7 (convergent transfer) to BAMC-1-bearing athymic mice, which were treated similarly with OK-432. More than 85% of the recipient athymic mice survived when an adoptive transfer was made on and after day 7. These results indicated that the effector cells developed before day 8 in euthymic mice. The effector cells detectable on day 7 in the PEC represent plastic- or nylon-wool-column-nonadherent cells, which could cure the tumor-bearing athymic mice. Furthermore, the effector cells were destroyed when the nylon-wool-column-nonadherent cells were treated with an anti-L3T4 antibody and complement whereas the same treatment with anti-Lyt2 antibody had no effect. These L3T4⁺ cells did not possess asialo-GM1 antigen. Although the exact mechanism of action of the effector cells is yet to be clarified, the induction of human equivalents of this type of effector cell would be a good parameter indicative of clinical effects induced by OK-432 or other biological response modifiers in an individual cancer patient.     

Protection of OK-432, a Streptococcus pyogenes preparation, against lethal infection of mice with herpes simplex virus

We have studied the protective effect of OK-432, a biological response modifier (BRM) of Streptococcus pyogenes origin, on the lethal infection of mice with herpes simplex virus (HSV)-1. A single intraperitoneal (i.p.) injection of more than 10 micrograms of OK-432, when given at least two days before the infection, gave a marked effect yielding nearly 100% protection against ordinarily lethal infection. The protection was independent of the amount of infected virus inoculated. When given after the infection, the agent even at the maximal dose (100 micrograms), produced only a marginal effect. A single i.p. administration of OK-432 augmented the natural killer (NK) activity of peritoneal exudate cells and spleen mononuclear cells in mice 2 to 3 days after injection of OK-432, coinciding with the times when it induced a survival effect on HSV-infection. Treating OK-432-treated mice with a combination of an anti-macrophage agent, silica, and an anti-NK cell agent, anti-asialo GM1 serum, before infection diminished the antiviral effect of OK-432. The OK-432 protection against HSV infection was also markedly diminished in athymic nude mice. Thus, the protective effect of OK-432 on lethal HSV infection seems to be based on the activation of NK cells, macrophages, and T lymphocytes.     

Mechanism of immunotherapeutic activity of OK-432 in the treatment of peritoneal carcinomatosis

Summary In this report the mechanism of therapeutic activity of OK-432 for the treatment of peritoneal carcinomatosis was investigated by correlating effector-cell augmentation with therapeutic activity in rats bearing MADB-106 carcinomatosis. Tumor cells were injected i.p. and the treatment with OK-432 was initiated 5 days later with 0.5, 1, 5 or 10 KE/animal of OK-432 injected i.p. semiweekly. Significant therapeutic activity was observed at all doses examined with greater prolongation of survival noted at the higher doses of OK-432. Animals treated with 0.5 KE/animal had a prolongation of the median survival time from 14 days for saline-treated animals to 17 days for the OK-432 treated animals (P<0.0008), while animals treated with higher doses had much longer periods of survival, some animals being tumor-free at 185 days. In the same studies, natural killer (NK) cell, lymphokine-activated killer cell, cytotoxic T lymphocyte, and macrophage tumoricidal/cytostatic activities were measured 7 days and 14 days following tumor injection (2 days and 9 days after initiation of immunotherapy). OK-432 had immunostimulatory activity in most of the assays of immune function examined and this correlated with host survival, including augmentation of peritoneal and peripheral blood cytotoxic T lymphocyte activity on day 14, peritoneal and alveolar macrophage activity on day 7 and day 14, as well as natural killer cell activity on day 14. These results suggest that the therapeutic doses are also immunomodulatory doses for the effector cells mentioned above. We suggest, therefore, that immunological monitoring may help to optimize treatment protocols for the treatment of peritoneal and perhaps pleural effusions with OK-432.By acceptance of this article, the publisher or recipient acknowledges the right of the U.S. Government to retain a nonexclusive, royalty-free license in and to any copyright covering the article. This research was sponsored by the National Cancer Institute, DHHS, under contract no. N01-23910 with Program Resources Inc. The contents of this publication do not necessarily reflect the views or policies of the Department of Health and Human Services, nor does mention of trade names, commercial products, or organizations imply endorsement by the U.S. Government     

The neutrophil as an information transmitter in tumor inhibition by a streptococcal biological response modifier,OK-432

 Effective treatment of a rat transplanted ascites tumor by i. p. injection of a streptococcal biological response modifier, OK-432, was abrogated by selective in vivo depletion of neutrophils by a monoclonal antibody, RP-3. The mechanisms by which neutrophils participate in the therapeutic action of OK-432 were studied with Winn’s assay using peritoneal exudate cells periodically obtained from rats i. p. injected with this biological response modifier. Intraperitoneal resident macrophages were first activated with OK-432, and within 3 h, tumor-inhibitory activity had moved to the early exuded neutrophils. However, 6 h after injection, exuded macrophages were the only cells involved in tumor inhibition. Considered together with other findings, it is likely that, in this system, neutrophils may transmit information from resident macrophages to exuded inflammatory macrophages in a series of responses induced by i. p. injection of OK-432. Received: 29 April 1996 / Accepted: 27 July 1996     

Antitumor effect of the streptococcal preparation OK-432 in a murine model of ovarian cancer

Summary The antitumor effects of the streptococcal preparation OK-432 were analyzed in a murine ovarian teratocarcinoma (MOT) model. Administration of OK-432 i.p. prevented tumor outgrowth in 75% of mice challenged with 10³ MOT cells i.p. 24 h previously. Treatment was less successful in mice challenged with 10⁴ or 10⁵ cells, preventing tumor growth in 25% of the former and only 5% of the latter group. Tumor-challenged mice cured by injections of OK-432 were not rendered resistant to a subsequent challenge with 10³ MOT cells 75 days after initial treatment. Only the i.p. route of administration was effective as i.v. OK-432 did not prolong survival of tumor-challenged mice. An antitumor response was detected as early as 24 h after i.p. treatment. This correlated temporally with an influx of neutrophils into the peritoneal cavity. Peritoneal cells obtained between 6 and 24 h after treatment were capable of lysing MOT targets in vitro. A single cell cytotoxicity assay demonstrated that peritoneal neutrophils, elicited by i.p. injection of OK-432, could bind to and lyse MOT targets. These data indicate that OK-432 is effective against small tumor cell inocula in this murine model of ovarian cancer and, furthermore, that the neutrophilic response into the peritoneal cavity plays a role in tumor rejection.     

Immunohistochemical analysis of inflammatory infiltrates around the injection site of a streptococcal preparation,OK-432, in rat tongue

On the assumption that neutrophils around the injection site of OK-432, a heat- and penicillin-treated lyophilized preparation of the Su strain ofStreptococcus pyogenes, enhance immunologic response through the production of Interleukin-1 (IL-1), OK-432 was injected into rat tongue, and specimens from the tongue were immunohistochemically investigated at various intervals after the injection, to clarify the process of inflammatory and immune responses at the injection site. Neutrophils and mononuclear cells appeared around the OK-432 injection site after 1 hour, increased to their maximum level at 24 hours, and then decreased from the 3rd to the 7th day. IL-1 was detected on neutrophils 3 hours after the injection, and OX-08-positive cells (suppressor/cytotoxic T cells and the majority of natural killer cells) remarkably increased. OX-39-positive cells (IL-2 receptor) appeared after 12 hours. These results suggest that neutrophils around the injection site of OK-432 at early phases of inflammation play a role in the expression of BRM function through IL-1.     

Augumentation of splenic antitumor immunity by local immunotherapy in gastric cancer patients

We previously reported that the antitumor effect of OK-432, a streptococcal preparation, was markedly augmented when this agent was injected into tumors together with fibrinogen. In order to elucidate the effect of this treatment on the spleen, we assessed splenic function in gastric cancer patients receiving preoperative local immunotherapy with OK-432 and fibrinogen. Immunohistochemical studies of the spleen at 7 days after intratumoral injection therapy revealed numerous macrophages phagocytizing OK-432 in the splenic sinuses. Phenotypic analysis of splenocytes by flow cytometry revealed an increase in the CD4/CD8 ratio and in the expression of HLA-DR, CD25, and Leu M3 by splenic T cells of the patients treated with OK-432 plus fibrinogen when compared to patients treated with OK-432 alone or untreated patients. Splenic T cells from patients treated with OK-432 plus fibrinogen showed significantly higher cytotoxicity against Daudi and K562 cells than T cells from control patients (p<0.05), and culture of these splenic T cells with recombinant IL-2 induced the expansion of lymphokine-activated killer cells. These results demonstrate that local immunotherapy with a mixture of OK-432 and fibrinogen effectively augumented splenic antitumor immunity in gastric cancer patients.     

Orally administered streptococcal preparation,OK-432 augments the antitumor immunity of patients with gastric or colorectal cancer

A streptococcal preparation, OK-432, was orally administered at a dose of 5 KE to patients with gastric or colorectal cancer for 7–14 days before their operations, and its immunomodulatory effects on peripheral blood lymphocytes (PBL), regional node lymphocytes (RNL) and tumor infiltrating lymphocytes (TIL) were assessed. The group treated with OK-432 included 8 gastric and 6 colorectal cancer patients, and the control group included 8 gastric and 8 colorectal cancer patients. The NK cell activity of PBL was significantly augmented by the oral administration of OK-432, and the proportions of Leu 7+ and Leu 11+ cells in PBL also increased. The responses of PBL and TIL to autologous tumor extracts in the presence of interleukin-2 were enhanced after the oral administration of OK-432. The proportion of OKT8+ cells in PBL increased after treatment with oral OK-432, whereas the proportion in RNL significantly decreased. These results indicate that oral OK-432 affects NK and T cells and may augment the antitumor immunity of patients with gastrointestinal cancer.     

Studies of the properties of a streptococcal preparation, OK-432 (NSC-B116209), as an immunopotentiator

Summary The present study was designed to examine the mechanism by which OK-432 triggers the cytotoxic activity of peritoneal exudate cells (PEC). When OK-432 was incubated with freshly harvested mouse serum, the formation of complexes of OK-432 with the third component of complement (C3) was demonstrated by using ¹³¹I-labeled mouse C3. The formation of C3-OK-432 complexes was totally abolished by a chelating compound, EDTA, which had been shown to inhibit the OK-432 induced activation of the alternative complement pathway. The C3-OK-432 complexes thus obtained bound to the resident PEC, which were subsequently shown to be activated. These activated PEC had augmented cytostatic activity against MM2 cells, a mouse mammary carcinoma.Further, the PEC from mice which had received an IP injection of OK-432 4–5 days previously were cytostatic against MM2 cells and also inhibited the growth of MM2 cells in culture. In contrast, resident PEC stimulated rather than inhibited the ³H-thymidine uptake by MM2 cells and the growth of MM2 cells. The mechanism of PEC (presumably macrophages) activation by OK-432 is discussed.     

Pronounced antitumor effect of LAK-like cells induced in the peritoneal cavity of mice after intraperitoneal injection of OK-432, a killed Streptococcal preparation

Summary More than 80% of BALB/c mice bearing BAMC-1 ascites tumor were completely cured after five consecutive (once every 2 days) i. p. injections of a 0.1 mg dose of OK-432, beginning on day 2 after tumor implantation. The antitumor effect of OK-432 was abolished in athymic nu/nu mice and in anti-thymocyte globulin-treated euthymic BALB/c mice, so although OK-432 treatment did increase the length of survival, all animals eventually died as a result of tumor growth. When peritoneal exudate cells (PEC), obtained on day 12 from OK-432-treated BAMC-1-bearing euthymic mice were evaluated for in vivo tumor neutralization activity, all mice receiving an i. p. injection of the admixture of the nonadherent PEC (1×10⁷ cells) with BAMC-1 cells (1×10⁵) survived for more than 60 days. When the same nonadherent PEC (1×10⁷ cells) were i. p. transferred adoptively 1 day after the inoculation of 1×10⁵ BAMC-1 tumor cells, again all mice survived.When these in vivo active PEC were tested for cytotoxicity in vitro against fresh BAMC-1 tumor cells, natural killer (NK) sensitive syngeneic RL 1, NK-sensitive allogeneic YAC-1 cells, NK-resistant syngeneic Meth-A cells, allogeneic tumor cells (EL4, B16, and P815) and xenogenic human cells, the PEC were found to be capable of lysing BAMC-1 tumor cells together with almost all of the other tumor cells, including NK-resistant cells. Nonadherent PEC contained at least two subpopulations of killer cells. One, directed to syngeneic BAMC-1 cells, was both Thy1.2 and asialo GM1 positive, and another, directed to allogeneic YAC-1 cells, was asialo GM1 positive but Thy1.2 negative. A cold target inhibition assay also suggested the presence of more than two subpopulations.These results indicate that T cells play a determined role in the immunotherapeutic effect of OK-432 on BALB/c mice bearing BAMC-1 tumor, although the participation of activated macrophages could not be excluded. The cells responsible for killing BAMC-1 and other tumor cells appearing in the PEC on day 12 were characterized as containing at least two kinds of lymphokine-activated killer cells.     

Peri-operative immunotherapy with OK-432

Pre- and postoperative intradermal administration of OK-432 enhanced the SU-PS skin reaction in patients with gastric cancer, but failed to prevent a fall in the NK activity induced by the operation.The change in NK activity was not associated with a change in the proportion of Leu 7-positive cells, but was related to Leu 11a-positive cells. Intradermal injection of OK-432 increased the proportion of Leu 7-positive cells in the patients in whom they accounted for less than 20% of lymphocyte population. The case was the same with Leu 11a-positive cells.Intravenous injection of OK-432 tended to increase suppressor-inducer T cells (CD4⁺2HA⁺ cells), B cells and Leu 7-positive cells. Particularly, the proportions of OK-M₁-positive cells and MHC class II antigen-positive cells increased in all patients. Immunotherapy with OK-432 given intravenously at a dose of 0.1 KE appeared to be safe because no side effects were essentially observed.     

Augmented induction of antitumor cells in vivo by cyclophosphamide fails to benefit antitumor resistance of the host

Summary The present study was designed to examine whether cyclophosphamide augmented induction of antitumor cells and antitumor resistance in C57BL/6 mice pretreated with mitomycin-C-treated EL4 cells (EL4_MMC) plus OK-432, a streptococcal preparation. C57BL/6 mice were pretreated with EL4_MMC (10⁷) plus OK-432 (2.5 KE) i.p. twice at 1-week intervals. When the mice received an i.p. injection of cyclophosphamide at 200 mg/kg 2 days before the last treatment, the antitumor activity of their spleen cells and peritoneal exudate cells (PEC) was effectively augmented 7–8 days after the last treatment. Splenic antitumor activity disappeared 15 days after the last treatment whereas augmented antitumor activity of the PEC was detected even 28 days after the last treatment. This cyclophosphamide effect was dose-dependent and 200 mg/kg was the most effective among the doses tested. If the EL4_MMC plus OK-432 treatment was injected at a s.c. site, it was also effective in combination with cyclophosphamide. The antitumor activity of the PEC from s.c.-pretreated mice, however, was lower than that from i.p.-pretreated mice. Despite the fact that cyclophosphamide effectively augmented induction of antitumor cells in C57BL/6 mice pretreated with EL4_MMC plus OK-432, it diminished rather than augmented, under all conditions tested, the ability of the mice to resist a challenge of live EL4 cells. Reduction of antitumor resistance by cyclophosphamide was also observed in an experimental system of a semi-syngeneic host (BDF₁) tumor (EL4). These results indicate that augmentation of in vivo induction of certain kinds of antitumor cells does not necessarily result in a beneficial augmentation of the host's ability to resist tumor growth.     

Stimulation of arachidonic acid metabolism by a streptococcal preparation (OK-432) in rat peritoneal macrophages.

A streptococcal preparation OK-432 is reported to be an immunopotentiator and a potent antitumor agent. In order to elucidate the mechanism of biologic action, effects of OK-432 on arachidonic acid metabolism in rat peritoneal macrophages were investigated. Prostaglandin E2 production and release of radioactivity from [3H]arachidonic acid-labeled macrophages were found to be stimulated by OK-432 in a concentration-dependent manner (5 to 80 micrograms/ml). Heat-treatment of OK-432 further stimulated its effects. These stimulative effects on arachidonic acid metabolism by OK-432 were not observed in MDCK cells that have no phagocytotic activity. Furthermore, cytochalasin B treatment completely suppressed the stimulative effects induced by OK-432 in macrophages. These results strongly indicate that the stimulative effects by OK-432 on arachidonic acid metabolism are dependent on phagocytosis of OK-432 particles. Significance of stimulation of arachidonic acid metabolism in macrophages by OK-432 for its biological effects is discussed.     

In vitro production of immune interferon (IFN gamma) by murine spleen cells when different sensitizing antigens are used in vivo and in vitro

OK-432, a killed preparation of Streptococcus pyogenes, as well as bacillus Calmette-Guérin (BCG) and Corynebacterium parvum are all known to induce immune interferon (IFN gamma) in mice. To examine the mechanisms of IFN gamma induction by OK-432, DDI mice were sensitized with various doses of OK-432, either by a single injection of a 1-mg dose or repeated injections of 0.1-mg doses given intraperitoneally. Spleen cells removed from the mice 7-9 days after the last injection produced high-titered IFN gamma (600-800 IU/ml) in vitro in the presence of 5 micrograms/ml of OK-432. In the absence of OK-432, however, in vitro IFN gamma production of sensitized spleen cells was quite limited. Moreover, when inducers of different antigenic entities such as serologically unrelated Streptococcus faecalis, Listeria monocytogenes, or Con A were added in vitro, instead of OK-432, to the OK-432-sensitized spleen cells, high-titered IFN gamma production also occurred. This may indicate that the signal required by T cells to produce IFN gamma in vitro need not necessarily be the same as that required to sensitize mouse macrophage in vivo. Once sensitized with OK-432, mice spleen cells continued to produce high-titered IFN gamma for more than 3 but less than 5 weeks.     

Therapeutic efficacy of sequential therapy with OK-432, cyclophosphamide,IL2-cultured lymphocytes andin vivo IL2 against advanced murine plasmacytoma

BALB/c mice inoculated IP with a syngeneic plasmacytoma MOPC104E were treated with a combination of a streptococcal preparation, OK-432 (1 KE, 0.1 mg/mouse), low-dose of cyclophosphamide (CPA, 1 mg/kg) and adoptive transfer of tumor-bearer-spleen cells (2 x 10(7) cells) cultured with IL2 and sonicated tumor extract (adoptive immunotherapy; AIT). The consecutive protocol of OK-432 (day 8, 9 post inoculation) - CPA (day 10) - AIT (day 11) was the most effective. Rate of complete remission was highest when recombinant (r-) IL2 was injected to the mice after AIT. Moreover, another bacterial preparation, Nocardia rubra cell wall skeleton and another low-dose chemotherapy, Mitomycin C could be used successfully instead of OK-432 or CPA. Transfer test of intraperitoneal cells (tumor cells plus host cells) of mice on day 11 post inoculation (on the day of AIT) revealed that OK-432 augmented the susceptibility of peritoneal cells to cultured lymphocytes in inhibition of transplantability, and that CPA after OK-432 augmented the anti-tumor effect of tumor-bearer-spleen cells which act synergistically with cultured lymphocytes. This therapy schedule seems to be the best model to augment the effect of AIT with minimal side effect.     

Streptococcal preparation OK-432: a new maturation factor of monocyte-derived dendritic cells for clinical use

For vaccinations based on dendritic cells (DCs), maturation of DCs is critical to the induction of T-cell responses. We tested the efficacy of streptococcal preparation OK-432 as a Good Manufacturing Practice (GMP)-grade maturation agent. OK-432 is currently used in Japan as a cancer immunotherapy drug. Immature monocyte-derived dendritic cells (imMo-DCs) isolated from human peripheral blood monocytes stimulated with granulocyte-macrophage colony stimulating factor and interleukin-4 were exposed to maturation factors, i.e., lipopolysaccharide (LPS), tumor necrosis factor alpha (TNF-alpha) plus prostaglandin E2 (PGE2), and OK-432 for 2 days. OK-432 increased expression of activation- and maturation-related molecules such as HLA-DR, CD80, CD83, and CD86 in imMo-DCs at levels similar to that of TNF-alpha plus PGE2, and higher than that of LPS. All agents examined induced allogeneic T-cell proliferation at a similar level. Only OK-432 caused significant production of interleukin-12 (IL-12) p70 and interferon gamma (IFN-gamma) at both the mRNA and protein levels in imMo-DCs. Neutralizing antibody against IL-12 p70 blocked IFN-gamma secretion from OK-432-stimulated Mo-DCs. IL-12 p70 produced by OK-432-stimulated imMo-DCs induced secretion of IFN-gamma by CD4+ T cells. OK-432 and LPS activated nuclear factor kappa B (NF-kappaB) in imMo-DCs. Both secretion of IL-12 p70 and IFN-gamma and activation of NF-kappaB induced by OK-432 were suppressed when imMo-DCs were pretreated with cytochalasin B. These results indicate that uptake of OK-432 by imMo-DCs is an early critical event for IL-12 p70 production and that NF-kappaB activation induced by OK-432 also contributes partially to IL-12 p70 production. In conclusion, OK-432 is a GMP-grade maturation agent and may be a potential tool for DC-based vaccine therapies.     

Induction of interferon-gamma in mouse spleen cells by OK-432, a preparation of Streptococcus pyogenes

A bacterial antitumor and immunopotentiating agent, OK-432, induced Interferon in the spleen cell cultures but not in the thymus cell cultures of various inbred strains of mice. When 1 × 10⁷ spleen cells were cultured in the presence of 5 μg/ml of OK-432, interferon activity was detected as early as 4 hr later and reached a maximum level of about 160 to 500 units/ ml 24 hr later. OK-432-induced interferon was mainly an IFN-γ of molecular weight approximately 40,000, but also contained IFN-α and IFN-β.     

Combined effects of cis-DDP and OK-432 on ascitic mastocytoma in mice: optimal period for the administration of OK-432

Combined effects of cis-DDP and OK-432 on the ascites mastocytoma was studied in mice laying emphasis on the best timing for the administration of the OK-432. Mice were transplanted with FMA3 mastocytoma cells into the abdominal cavity at a dose of 10(5) cells per mouse. They were injected with cis-DDP next day at a single shot of 8 mg/kg in the abdominal cavity. A streptococcal preparation, OK-432, was i.p. injected two times at intervals of one week at a dose of 50 KE/kg per injection. The pair of injections started 1, 2, 3, 4 or 5 weeks after the transplantation. Mean survival time (M. S. T.) of mice was 16.8, 16.2 and 52.3 in the groups of mice nontreated, given OK-432 alone, and given cis-DDP alone, respectively. In comparison of M. S. T. within the groups of mice treated with the combination of cis-DDP and OK-432, the highest value was observed in the group which was given OK-432 between 3 and 4 weeks after the transplantation.